Pioneering Mammalian Embryo Culture

  • John D. Biggers


In the fast pace and competitiveness of modern science, there is less and less time to teach students the background on how the currently used scientific methods and ideas came about. This fact is unfprtunate, for it is salutary to realize that many so-called discoveries and inventions are not new and that the old literature is replete with ideas. In doing our work we often stand on the shoulders of those who have gone before. The study of early mammalian development in vitro, which has undergone explosive growth since 1960, provides many illustrations of these facts. In addition, to minimize the tendency of investigators to “rediscover the wheel”, it is pertinent at this time to reflect on the roots of the field. The subject is closely intertwined with the history of embryo transfer (see Adams, 1982) and of in vitro fertilization [see Austin (1961) and Biggers (1984)].


Mouse Embryo Embryo Transfer Preimplantation Embryo Amino Acid Requirement Mouse Blastocyst 
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  1. Adams, C.E., 1956, Egg transfer and fertility in the rabbit, Proc. Int. Congr. Anim. Reprod. (Cambridge), Section III, pp. 5-6.Google Scholar
  2. Adams, C.E., 1965, The influence of maternal environment on preimplantation stages of pregnancy in the rabbit, in: Preimplantation Stages of Pregnancy (G.E.W. Wolstenholme, and M. O’Connor, eds.), Churchill, London, pp. 345–377.Google Scholar
  3. Adams, C.E., 1970, The development of rabbit eggs after culture in vitro for 1–4 days, J. Embryol. Exp. Morphol. 23: 21–34.PubMedGoogle Scholar
  4. Adams, C.E., 1982, Egg transfer: historical aspects, in: Mammalian Egg Transfer (C.E. Adams, ed.), CRC Press, Boca Raton, FL, pp. 1–18.Google Scholar
  5. Austin, C.R., 1961, The Mammalian Egg, Charles C. Thomas, Springfield, IL.Google Scholar
  6. Biggers, J.D., 1984, In vitro fertilization and embryo transfer in historical perspective, in: In Vitro Fertilization and Embryo Transfer (A. Trounson, and C. Wood, eds.), Churchill Livingstone, London, pp. 3–15.Google Scholar
  7. Biggers, J.D., and Brinster, R.L., 1965, Biometrical problems in the study of early mammalian embryos in vitro, J. Exp. Zool. 158: 39–47.PubMedGoogle Scholar
  8. Biggers, J.D., and McLaren, A., 1958, “Test tube” animals. The culture and transfer of early mammalian embryos, Discovery (London) 19: 423–426.Google Scholar
  9. Biggers, J.D., Rinaldini, L.M., and Webb, M., 1957, The study of growth factors in tissue culture, Symp. Soc. Exp. Biol. 11: 264–297.PubMedGoogle Scholar
  10. Biggers, J.D., Gwatkin, R.B.L., and Brinster, R.L., 1962, Development of mouse embryos in organ cultures of fallopian tubes on a chemically defined medium, Nature (London) 194: 747–749.Google Scholar
  11. Biggers, J.D., Moore, B.D., and Whittingham, D.G., 1965, Development of mouse embryos in vivo after cultivation from two-cell ova to blastocysts in vitro, Nature (London) 206: 734–735.Google Scholar
  12. Biggers, J.D., Whittingham, D.G., and Donahue, R.P., 1967, The pattern of energy metabolism in the mouse oocyte and zygote, Proc. Natl. Acad. Sci. USA 58: 560–567.PubMedGoogle Scholar
  13. Biggers, J.D., Whitten, W.K., and Whittingham, D.G., 1971, The culture of mouse embryos in vitro, in: Methods in Mammalian Embryology (J.C. Daniel, Jr., ed.), Freeman, San Francisco, pp. 86–116.Google Scholar
  14. Brachet, A., 1912, Développement in vitro de blastodermes et de jeunes embryons de mammifères, C.R. Hebd. Seances Acad. Sci. 155: 1191–1193.Google Scholar
  15. Brachet, A., 1913, Recherches sur le déterminisme héréditaire de l’oeuf des mammifères. Développement in vitro de jeunes vésicules blastodermiques du lapin, Arch. Biol. (Paris) 28: 447–503.Google Scholar
  16. Brinster, R.L., 1963, A method for in vitro cultivation of mouse ova from two-cell to blastocyst, Exp. Cell Res. 32: 205–207.PubMedGoogle Scholar
  17. Brinster, R.L., 1965a, Studies on the development of mouse embryos in vitro. I. The effect of osmolality and hydrogen ion concentration, J. Exp. Zool. 158: 49–57.PubMedGoogle Scholar
  18. Brinster, R.L., 1965b, Studies on the development of mouse embryos in vitro. II. The effect of energy sources, J. Exp. Zool. 158: 59–68.PubMedGoogle Scholar
  19. Brinster, R.L., 1965c, Studies on the development of mouse embryos in vitro. IV. Interaction of energy sources, J. Reprod. Fertil. 10: 227–240.PubMedGoogle Scholar
  20. Brinster, R.L., 1970, Culture of two-cell rabbit embryos to morulae, J. Reprod. Fertil. 21: 17–22.PubMedGoogle Scholar
  21. Brinster, R.L., 1971, In vitro culture of the embryo, in: Pathways to Conception (A.I. Sherman, ed.), Charles C. Thomas, Springfield, IL, pp. 245–277.Google Scholar
  22. Castle, W.E., and Gregory, P.W., 1929, The embryological basis of size inheritance in the rabbit, J. Morphol. 48: 81–103.Google Scholar
  23. Chang, M.C., 1947, Normal development of fertilized rabbit ova stored at low temperature for several days, Nature (London) 159: 602–603.Google Scholar
  24. Chang, M.C., 1948, Transplantation of fertilized rabbit ova: the effect on viability of age, in vitro storage period, and storage temperature, Nature (London) 161: 978–979.Google Scholar
  25. Charlton, H.H., 1917, The fate of the unfertilized egg in the white mouse, Biol. Bull. 33: 321–338.Google Scholar
  26. Cole, R.J., Edwards, R.G., and Paul, J., 1964, Cytodifferentiation in cell colonies and cell strains derived from cleaving ova and blastocysts of the rabbit, Exp. Cell Res. 37: 501–504.Google Scholar
  27. Cole, R.J., and Paul, J., 1965, Properties of cultured preimplantation mouse and rabbit embryos, and cell strains derived from them, in: Preimplantation Stages of Pregnancy (G.E.W. Wolstenholme, and M. O’Connor, eds.), Churchill, London, pp. 82–122.Google Scholar
  28. Daniel, J.C., Jr., 1964, Some effects of steroids on cleavage of rabbit eggs in vitro, Endocrinology 75: 706–710.PubMedGoogle Scholar
  29. Daniel, J.C., Jr., 1965, Studies on the growth of 5-day old rabbit blastocysts in vitro, J. Embryol. Exp. Morphol. 13: 83–95.PubMedGoogle Scholar
  30. Daniel, J.C., Jr., and Krishnan, R.S., 1967, Amino acid requirements for growth of the rabbit blastocyst in vitro, J. Cell. Comp. Physiol. 70: 155–160.Google Scholar
  31. Daniel, J.C., Jr., and Levy, J.D., 1964, Action of progesterone as a cleavage inhibitor of rabbit ova in vitro, J. Reprod. Fertil. 7: 323–329.PubMedGoogle Scholar
  32. Daniel, J.C., Jr., and Olson, J.D., 1968, Amino acid requirements for cleavage of the rabbit ovum, J. Reprod. Fertil. 15: 453–455.PubMedGoogle Scholar
  33. Defrise, A., 1933, Some observations on living eggs and blastulae of the albino rat, Anat. Rec. 57: 239–250.Google Scholar
  34. Ducibella, T., and Anderson, E., 1975, Cell shape and membrane changes in the eight-cell mouse embryo: prerequisites for morphogenesis of the blastocyst, Dev. Biol. 45: 231–250.PubMedGoogle Scholar
  35. Eagle, H., 1959, Amino acid metabolism in mammalian cell cultures, Science 130: 432–437.PubMedGoogle Scholar
  36. Earle, W.R., 1943, Production of malignancy in vitro. IV. The mouse fibroblast cultures and changes seen in the living cells, J. Natl. Cancer Inst. 4: 165–212.Google Scholar
  37. Edwards, R.G., 1964, Cleavage of one-and two-celled rabbit eggs in vitro after removal of the zona pellucida, J. Reprod. Fertil. 7: 413–415.PubMedGoogle Scholar
  38. Epstein, C.J., Smith, S., Travis, B., and Tucker, G., 1978, Both X-chromosomes function before visible X-chromosome inactivation in female mouse embryos, Nature (London) 274: 500–503.Google Scholar
  39. Fischer, A., 1947, Biology of Tissue Cells, Cambridge University Press, Cambridge, England.Google Scholar
  40. Gardner, R.L., 1968, Mouse chimeras obtained by the injection of cells into the blastocyst, Nature (London) 220: 596–597.Google Scholar
  41. Gardner, R.L., 1985a, Origin and development of the trophectoderm and inner cell mass, in: Implantation of the Human Embryo (R.G. Edwards, J.M. Purdy, and P.C. Steptoe, eds.), Academic Press, London, pp. 155–178.Google Scholar
  42. Gardner, R.L., 1985b, Clonal analysis of early mammalian development, Phil. Trans. Roy. Soc. Lond. B 312: 163–178.Google Scholar
  43. Gardner, R.L., and Edwards, R.G., 1968, Control of the sex ratio at full term in the rabbit by transferring sexed blastocysts, Nature (London) 218: 346–348.Google Scholar
  44. Gardner, R.L., and Johnson, M.H., 1973, Investigation of early mammalian development using interspecific chimaeras between rat and mouse, Nature (London) New Biol. 246: 86–89.Google Scholar
  45. Gardner, R.L., Papaioannou, V.E., and Barton, S.C., 1973, Origin of the ectoplacental cone and secondary giant cells in mouse blastocysts reconstituted from isolated trophoblast and inner cell mass, J. Embryol. Exp. Morphol. 30: 561–572.PubMedGoogle Scholar
  46. Gregory, P.W., 1930, The early embryology of the rabbit, Carnegie Inst. Wash. Publ. 21: 141–168.Google Scholar
  47. Gregory, P.W., and Castle, W.E., 1931, Further studies on the embryological basis of size inheritance in the rabbit, J. Exp. Zool. 59: 199–211.Google Scholar
  48. Gwatkin, R.B.L., 1963, Effect of viruses on early mammalian development, I. Action of Mengo encephalitis virus on mouse ova cultivated in vitro, Proc. Natl. Acad. Sci. USA 50: 576–581.PubMedGoogle Scholar
  49. Gwatkin, R.B.L., 1966a, Defined media and development of mammalian eggs in vitro, Ann. N.Y. Acad. Sci. 139: 79–90.PubMedGoogle Scholar
  50. Gwatkin, R.B.L., 1966b, Amino acid requirements for attachment and outgrowth of the mouse blastocyst in vitro, J. Cell Physiol. 68: 335–344.Google Scholar
  51. Gwatkin, R.B.L., and Meckley, P.E., 1966, Chromosomes of the mouse blastocyst following its attachment and outgrowth in vitro, Ann. Med. Exp. Biol. Fenn. 44: 125–127.PubMedGoogle Scholar
  52. Ham, R.G., 1963, An improved nutrient solution for diploid Chinese hamster and human cell lines, Exp. Cell Res. 29: 515–526.PubMedGoogle Scholar
  53. Hammett, F.S., 1930, The natural chemical regulation of growth by increase in cell number, Proc. Amer. Phil. Soc. 69: 217–223.Google Scholar
  54. Hammond, J., Jr., 1949, Recovery and culture of tubal mouse ova, Nature (London) 163: 28–29.Google Scholar
  55. Harrison, R.G., 1969, Organization and Development of the Embryo, Yale University Press, New Haven, CT, pp. 67–116.Google Scholar
  56. Heape, W., 1891, Preliminary note on the transplantation and growth of mammalian ova within a uterine foster-mother, Proc. Roy. Soc. Lond. B Biol. Sci. 48: 457–458.Google Scholar
  57. Heape, W., 1897, Further note on the transplantation and growth of mammalian ova within a uterine foster-mother, Proc. Roy. Soc. Lond. B Biol. Sci. 62: 178–183.Google Scholar
  58. Hsu, Y., 1971, Post-blastocyst differentiation in vitro, Nature (London) 231: 100–102.Google Scholar
  59. Kane, M.T., and Foote, R.H., 1970, Culture of two-and four-cell rabbit embryos to the expanding blastocyst stage in synthetic media, Proc. Soc. Exp. Biol. Med. 133: 921–925.PubMedGoogle Scholar
  60. Kane, M.T., and Foote, R.H., 1971, Factors affecting blastocyst expansion of rabbit zygotes and young embryos in defined media, Biol. Reprod. 4: 41–47.PubMedGoogle Scholar
  61. Krebs, H.A., and Henseleit, K., 1932, Untersuchungen über die Harnstoffbildung im Tierkorper, Z. Phys. Chem. 210: 33–66.Google Scholar
  62. Lewis, W.H., 1931, Living mouse eggs, Anat. Rec. 48: 52 (abs).Google Scholar
  63. Lewis, W.H., and Gregory, P.W., 1929, Cinematographs of living developing rabbit-eggs, Science 69: 226–229.PubMedGoogle Scholar
  64. Lewis, W.H., and Hartman, C.G., 1933, Early cleavage stages of the egg of the monkey (Macacas rhesus), Carnegie Inst. Wash. Publ. 24: 187–203.Google Scholar
  65. Long, J.A., and Evans, H.M., 1922, The oestrous cycle in the rat and its associated phenomena, Mem. Univ. Calif. 6: 1–148.Google Scholar
  66. Lwoff, A., Dulbecco, M., Vogt, M., and Lwoff, M., 1955, Kinetics of the release of poliomyelitis virus from single cells, Virology 1: 128–139.PubMedGoogle Scholar
  67. Mark, E.L, and Long, J.A., 1912, The living eggs of rats and mice with a description of apparatus for obtaining and observing them, Univ. Calif. Publ. Zool. 9: 105–136.Google Scholar
  68. Mather, W.B., 1950, The technique of rabbit blastoderm culture, Univ. Queensland Pap. Dept. Biol. 2 (No. 15): 1–8.Google Scholar
  69. Maurer, R.R., Whitener, R.H., and Foote, R.H., 1969, Relationship of in vivo gamete aging and exogenous hormones to early embryo development in rabbits, Proc. Soc. Exp. Biol. Med. 131: 882–885.PubMedGoogle Scholar
  70. Maurer, R.R., Onuma, H., and Foote, R.H., 1970, Viability of cultured and transferred rabbit embryos, J. Reprod. Fertil. 21: 417–422.PubMedGoogle Scholar
  71. Maximow, A., 1925, Tissue-cultures of young mammalian embryos, Carnegie Inst. Wash. Publ. 16: 47–113.Google Scholar
  72. McLaren, A., and Biggers, J.D., 1958, Successful development and birth of mice cultivated in vitro as early embryos, Nature (London) 182: 877–878.Google Scholar
  73. McLaren, A., and Michie, D., 1956, Studies in the transfer of fertilized mouse eggs to uterine foster mothers. I. Factors affecting the implantation and survival of native and transferred eggs, J. Exp. Biol. 33: 394–416.Google Scholar
  74. Miller, B.J., and Reimann, S.P., 1940, Effect of DL-methionine and L-cysteine on the cleavage rate of mammalian eggs, Arch. Pathol. 29: 181–188.Google Scholar
  75. Mintz, B., 1962a, Experimental study of the developing mammalian egg; removal of the zona pellucida, Science 138: 594.PubMedGoogle Scholar
  76. Mintz, B., 1962b, Experimental recombination of cells in the developing mouse egg. Normal and lethal mutant genotypes, Amer. Zool. 2: 145 (abs).Google Scholar
  77. Mintz, B., 1964, Formation of genetically mosaic mouse embryos, and early development of “lethal (t12/t12)-normal” mosaics, J. Exp. Zool. 157: 273–285.PubMedGoogle Scholar
  78. Naglee, D.L., Maurer, R.R., and Foote, R.H., 1969, Effect of osmolarity on in vitro development of rabbit embryos in a chemically defined medium, Exp. Cell Res. 58: 331–333.PubMedGoogle Scholar
  79. Onuma, H., Maurer, R.R., and Foote, R.H., 1968, in vitro culture of rabbit ova from early cleavage stages to the blastocyst stage, J. Reprod. Fertil. 16: 491–494.PubMedGoogle Scholar
  80. Pincus, G., 1927, Comparative study of the chromosomes of the Norway rat (Ratios norvegius ersel.) and the black rat (Rattus rattus L.), J. Morphol. 44: 515–540.Google Scholar
  81. Pincus, G., 1930, Observations on the living eggs of rabbits, Proc. Roy. Soc. Lond. B Biol. Sci. 107: 132–167.Google Scholar
  82. Pincus, G., 1936, The Eggs of Mammals, Macmillan, New York, pp. 1–160.Google Scholar
  83. Pincus, G., 1937, The metabolism of ovarian hormones, especially in relation to the growth of the fertilized ovum, Cold Spring Harbor Symp. 5: 44–56.Google Scholar
  84. Pincus, G., 1941, Factors controlling the growth of rabbit blastocysts, Amer. J. Physiol. 133: P412–P413.Google Scholar
  85. Pincus, G., and Werthessen, N.T., 1938, The comparative behaviour of mammalian eggs in vivo and in vitro, J. Exp. Zool. 78: 1–18.Google Scholar
  86. Purshottam, N., and Pincus, G., 1961, In vitro cultivation of mammalian eggs, Anat. Rec. 140: 51–55.PubMedGoogle Scholar
  87. Smith, A.U., 1949, Cultivation of rabbit eggs and cumuli for phase-contrast microscopy, Nature (London) 164: 1136–1137.Google Scholar
  88. Smith, A.U., 1952, Behaviour of fertilized rabbit eggs exposed to glycerol and to low temperatures, Nature (London) 170: 374–375.Google Scholar
  89. Smith, A.U., 1953, In vitro experiment with rabbit eggs, in: Mammalian Germ Cells (G.E.W. Wolstenholme, M.P. Cameron, and S.J. Freeman, eds.), Churchill, London, p. 217.Google Scholar
  90. Squier, R.R., 1932, The living egg and early stages of its development in the guinea-pig, Carnegie Inst. Wash. Publ. 21: 225–250.Google Scholar
  91. Staples, R.E., 1967, Development of 5-day rabbit blastocysts after culture at 37°C, J. Reprod. Fertil. 13: 369–372.PubMedGoogle Scholar
  92. Tarkowski, A.K., 1961, Mouse chimeras developed from fused eggs, Nature (London) 190: 857–860.Google Scholar
  93. Tarkowski, A.K., 1963, Studies on mouse chimeras developed from eggs fused in vitro, in: Symposium on Organ Culture, Natl. Cancer Inst. Monograph No. 11 (C.J. Dawe, ed.), National Cancer Institute, Bethesda, MD, pp. 51–70.Google Scholar
  94. Umbreit, W.W., Burris, R.H., and Stauffer, J.F., 1949, Manometric Techniques and Tissue Metabolism, Burgess, Minneapolis, MN, pp. 118–119.Google Scholar
  95. Waddington, C.H., and Waterman, A.J., 1933, The development in vitro of young rabbit embryos, J. Anat. 67: 356–370.Google Scholar
  96. Washburn, W.W., 1951, A study of the modifications in rat eggs observed in vitro and following tubal retention, Arch. Biol. 62: 439–458.Google Scholar
  97. Waterman, A.J., 1934, Survival of young rabbit embryos on artificial media, Proc. Natl. Acad. Sci. USA 20: 145–146.PubMedGoogle Scholar
  98. Waymouth, C., 1959, Rapid proliferation of sublines of NCTC clone 929 (Strain L) mouse cells in a simple chemically defined medium (MB752/1), J. Natl. Cancer Inst. 22: 1003–1016.PubMedGoogle Scholar
  99. Whitten, W.K., 1956a, Culture of tubal ova, Nature (London) 177: 96.Google Scholar
  100. Whitten, W.K., 1956b, Physiological control of population growth, Nature (London) 178: 992.Google Scholar
  101. Whitten, W.K., 1957a, Culture of tubal ova, Nature (London) 179: 1081–1082.Google Scholar
  102. Whitten, W.K., 1957b, The effect of progesterone on the development of mouse eggs in vitro, J. Endocrinol. 16: 80–85.PubMedGoogle Scholar
  103. Whitten, W.K., and Biggers, J.D., 1968, Complete development in vitro of the pre-implantation stages of the mouse in a simple chemically defined medium, J. Reprod. Fertil. 17: 399–401.PubMedGoogle Scholar
  104. Whittingham, D.G., 1968, Development of zygotes in cultured mouse oviducts. I. The effect of varying oviductal conditions, J. Exp. Zool. 169: 391–398.PubMedGoogle Scholar
  105. Whittingham, D.G., 1975, Fertilization, early development and storage of mammalian ova in vitro, in: The Early Development of Mammals (M. Balls, and A.E. Wild, eds.), Cambridge University Press, Cambridge, England, pp. 1–24.Google Scholar
  106. Whittingham, D.G., and Biggers, J.D., 1967, Fallopian tube and early cleavage in the mouse, Nature (London) 213: 942–943.Google Scholar
  107. Whittingham, D.G., Leibo, S.P., and Mazur, P., 1972, Survival of mouse embryos frozen to-196°C and-296°C, Science 113: 247.Google Scholar
  108. Wilmut, I., 1972, Effect of cooling rate, warming rate, cryoprotective agent and stage of development on survival of mouse embryos during cooling and thawing, Life Sci. 11 (pt. 2): 1071–1079.Google Scholar
  109. Wilson, I.B., Bolston, E., and Cuttler, R.H., 1972, Preimplantation differentiation in the mouse egg as revealed by microinjection of vital markers, J. Embryol. Exp. Morphol. 27: 467–479.PubMedGoogle Scholar
  110. Wolstenholme, G.E.W., and O’Connor, M., 1965, Preimplantation Stages of Pregnancy, Churchill, London.Google Scholar
  111. Ziomek, C.A., and Johnson, M.H., 1980, Cell surface interaction induces polarization of mouse 8-cell blastomeres at compaction, Cell 21: 935–942.PubMedGoogle Scholar

Copyright information

© Plenum Press, New York 1987

Authors and Affiliations

  • John D. Biggers
    • 1
  1. 1.Laboratory of Human Reproduction and Reproductive Biology, and Department of Physiology and BiophysicsHarvard Medical SchoolBostonUSA

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